Many applications require detection and identification of ligands, or molecules having particular binding properties. The binding properties of a particular ligand may be detected and characterized by the use of appropriate sensors. “Biosensors” have been reported in the literature and provide an alternative model for molecular screening. Biosensors are made up of an analytical platform and a binding entity to which the ligand may bind. The detection of a ligand by a biosensor requires a ligand which binds the binding entity and an analytical platform or sensor which generates a detectable signal that can be measured. The analytical platforms measure changes in mass, capacitance, resistance, surface plasma resonance, reflectometric interference, etc. resulting from the interaction of the ligand with the binding entity.
The current state of the art in biosensor technology includes a number of biosensor designs. For example, U.S. Pat. No. 6,241,863 (with inventor Monbouquette) describes the development of amperometric biosensors based on redox enzymes. U.S. Pat. No. 6,239,255 describes surface plasmon resonance biosensors. Still other biosensors have been described, including biosensors which utilize functionalized microspheres for optical diffraction (U.S. Pat. No. 6,221,579), mass-sensitive biosensors (U.S. Pat. No. 6,087,187), hybrid biosensors (U.S. Pat. No. 6,051,422), metal oxide matrix biosensors (U.S. Pat. No. 5,922,183), silicon-based biosensors (U.S. Pat. No. 5,874,047), solid-supported membrane biosensors (U.S. Pat. No. 5,846,814), fiber-optic chemiluminescent biosensors (U.S. Pat. No. 5,792,621) and others.
Biosensors previously reported in the literature are somewhat limited because the reported devices have low sensitivity, limited longevity, and/or long response times. Decker et al. ((2000) J. Immunol. Methods 233:159-165) reported that more than 90 minutes were needed to measure phage binding by peptide fragments immobilized by biotin/streptavidin coupling. Hengerer et al. ((1999) Biotechniques 26: 956-60, 962, 964) reported binding of phage antibodies to antigen immobilized on a quartz crystal microbalance with a time constant of about 100 min. These long response times are not compatible with rapid screening and make large-scale screening unwieldy. Therefore, there remains a need for a biosensor which can rapidly detect specific proteins.
In addition, reported biosensors generally suffer from disadvantages such as low specificity and low affinity. Some biosensor platforms utilize antibodies as the binding element. For example, U.S. Pat. No. 5,922,183 teaches the use of thin film composites of metal oxides and antibodies for amperometric and potentiometric sensing. Porous silicon biosensors are described for use with antibodies in U.S. Pat. No. 5,874,047. A patterned multiple antibody substrate for use in biosensors or immunosensors was prepared by adsorbing specific antibodies at the sites in U.S. Pat. No. 5,858,801. U.S. Pat. No. 5,039,611 teaches the use of monoclonal antibodies to superficial papillary bladder tumor cells in an ELISA-type format. See also, copending U.S. application Ser. No. 09/452,968, filed Dec. 2, 1999.
Antibody-based sensors represent an improvement over previously-used sensors in several ways, and can exhibit improved specificity and affinity (see, e.g., Ziegler et al. (1998) Biosensors & Bioelectronics 13: 539-571. However, antibody-based sensors have several disadvantages which restrict their usefulness, including high cost and short longevity or inability to perform in various environmental or field test conditions. Moreover, the quality of antibodies can vary with different production variables, such as the animal used to produce the antibodies. Another disadvantage of antibodies is that it may take months to generate the desired antibodies for use in an antibody-based sensor.
The threat of bioterrorism highlights the need for specific, accurate sensors that are rapidly prepared. At present, the earliest recognition of and response to a bioterrorist attack with Bacillus anthracis (anthrax) spores may be based on clinical manifestations of anthrax and laboratory culture tests, which require days to complete (Inglesby et al. (1999) JAMA 281: 1735-45). Thus, a need exists for specific, accurate biosensors that are rapidly prepared.